Figure 3 Immunohistochemistry Validation of BACE in Mouse Brain
Immunohistochemical analysis of paraffin-embedded mouse brain tissue using anti-BACE antibody (2253) at 2.5 μg/ml. Tissue was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4˚C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin. See high quality image.

Figure 5 Immunocytochemistry Validation of BACE in 3T3/NIH Cells
Immunohistochemical analysis of 3T3/NIH cells using anti-BACE antibody (2253) at 10 μg/ml. Cells was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4˚C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin. See high quality image.

Figure 6 KO and Overexpression Validation of BACE in Human and Mouse Brain and 293 Cells. (Singer et al., 2005)
Western blot analysis of the BACE1 (2253) antibody’s ability to recognize human and murine BACE1. The BACE1 antibody recognized both the mouse and human forms of BACE1. Lanes 1–4 are frontal cortex homogenates from human and mouse brains. Lane 1 is from a neurologically unimpaired aged human control case, lane 2 from a BACE1-deficient mouse, lane 3 from a nontransgenic mouse and lane 4 from hBACE1 transgenic mouse. Lanes 5–7 are lysates from HEK293T cells transfected with a plasmid vector expressing eGFP, mBACE1 and hBACE1, respectively. See high quality image.

Figure 7 KD Validation of BACE in Mouse Brain (Singer et al., 2005)
Characterization of the effects of lenti-siBACE1-6 expression in the brains of APP transgenic mice. (a–d) Anti-eGFP immunoreactivity in the hippocampus (the injection site) shows comparable and consistent expression of lenti-siRNA constructs in the dentate gyrus (dg) and stratus polymorphus (sp). (e) Anti-BACE1 immunoreactivity in the hippocampus of nontransgenic mice treated with lenti-siGlut4. (f) Reduced BACE1 immunostaining in the hippocampus of nontransgenic mice treated with lenti-siBACE1-6 vector. (g) Intense BACE1 immunoreactivity in the hippocampus of APP transgenic mice treated with lenti-siGlut4. (h) Reduced BACE1 expression in APP transgenic mice treated with lenti-siBACE1-6 vector. (i,j) Anti-BACE1 reacted with pyramidal cell bodies in the neocortex, which was not injected, See high quality image.

Figure 8 KD Validation of BACE in Mouse Brain (Singer et al., 2005)
Immunolabeling patterns of BACE1 expression and the lenti-siRNA distribution. Sections from APP transgenic mice treated with the eGFPtagged lenti siRNA vectors (green) were co-immunolabeled with an antibody against BACE1 (red) and imaged with the LSCM. All sections are from the hippocampus of treated mice. (a–c) Lenti-siBACE1-6–treated mice. Areas within the hippocampus expressing the eGFP tagged vector have reduced BACE1 immunolabeling. (d–f) Mice treated with the eGFP-tagged control lenti-siGlut4 show unchanged expression of BACE1 in the hippocampus. (g–i) Mice treated with a saline vehicle show unchanged expression of BACE1 in the hippocampus.. See high quality image.

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Specifications

SPECIES REACTIVITY:

Human

TESTED APPLICATIONS:

IHC, WB

APPLICATIONS:

These polyclonal antibodies can be used for detection of APP, BACE and BACE2 by immunoblot at 1 - 5 μgg/mL. APP and BACE antibodies can detect their respective proteins via immunohistochemistry at 2 - 10 μg/mL.

USER NOTE:

Optimal dilutions for each application to be determined by the researcher.

SPECIFICITY:

Both APP antibodies will react with the C99 fragment of APP.

IMMUNOGEN:

Rabbit polyclonal antibodies were raised against peptides corresponding to amino acid sequences from each of the corresponding proteins.

Optimal dilutions for each application to be determined by the researcher.

Background

BACKGROUND:

Accumulation of the amyloid-β peptide (Aβ) in the cerebral cortex is a critical event in the pathogenesis of Alzheimer’s disease. The βamyloid protein precursor (APP) is cleaved by one of two βsecretases (BACE and BACE2), producing a soluble derivative of the protein and a membrane anchored 99-amino acid carboxy-terminal fragment (C99). The C99 fragment serves as substrate for βsecretase to generate the 4 kDa amyloid-β peptide (Aβ), which is deposited in the Alzheimer’s disease patients’ brains. BACE was identified by several groups independently and designated β-site APP cleaving enzyme (BACE) . BACE is a transmembrane aspartic protease and co-localizes with APP. BACE2 also cleaves APP at β-site and at a different site within Aβ. BACE2 locates on chromosome 21q22.3, the so-called ‘Down critical region’, suggesting that BACE2 and Aβ may also contribute to the pathogenesis of Down syndrome.